Pulmonary infection with an exaggerated inflammatory response is the major cause of morbidity and mortality in cystic fibrosis (CF). The objective of this study was to determine whether differences in the innate immune system underlie the exaggerated immune response in CF. We established a model that recapitulates the exaggerated immune response in a CF mouse model by exposure to Pseudomonas aeruginosa LPS and assessed the pulmonary cellular and cytokine responses of wild-type (WT) and CF mice. Compared with WT mice, CF mice had increased numbers of neutrophils and increased proinflammatory cytokines in their bronchoalveolar lavage fluid after LPS exposure. Based on the increased levels of IL-1alpha, IL-6, granulocyte colony-stimulating factor (G-CSF), and keratinocyte chemoattractant, all of which are known to be produced by macrophages, we tested whether two populations of macrophages, bone marrow-derived macrophages and alveolar macrophages, directly contribute to the elevated cytokine response of CF mice to LPS. After in vitro stimulation of bone marrow-derived macrophages and alveolar macrophages with LPS, IL-1alpha, IL-6, G-CSF, and monocyte chemoattractant protein-1 were higher in CF compared with WT cell supernatants. Quantitative analyses for IL-6 and keratinocyte chemoattractant revealed that LPS-stimulated CF macrophages have higher mRNA and intracellular protein levels compared with WT macrophages. Our data support the hypothesis that macrophages play a role in the exuberant cytokine production and secretion that characterizes CF, suggesting that the macrophage response may be an important therapeutic target for decreasing the morbidity of CF lung disease.